Analysis of a Chinese pedigree affected with van der Woude syndrome due to variant of IRF6 gene / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic characteristics of a Chinese pedigree affected with van der Woude syndrome (VWS).@*METHODS@#A proband who had visited the Drum Tower Hospital Affiliated to Nanjing University Medical School in May 2020 for "two previous pregnancies with cleft lip and palate" was selected as the study subject. Trio-whole exome sequencing (trio-WES) was carried out for the patient. Candidate variants were verified by Sanger sequencing of her pedigree members (8 individuals from four generations) and bioinformatic analysis. Chromosomal microarray analysis (CMA) was used to rule out copy number variations in the fetuses.@*RESULTS@#Trio-WES revealed that the proband and her father had both harbored a heterozygous c.742G>T (p.G248C) missense variant of the IRF6 gene, for which her mother was of the wild type. The variant was located in a region with important functions and has not been reported previously. Prediction with several software suggested that it is likely to have a significant impact on the protein structure/function and is highly correlated with the specific phenotypes in this pedigree. Sanger sequencing confirmed co-segregation of the genotypes and phenotypes in the pedigree. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), this variant was rated as likely pathogenic (PM1+PM2_Supporting+PP1+PP3+PP4). Based on the above results, pre-implantation genetic diagnosis was carried out for the proband, which has led to birth of a healthy offspring with normal results for both site testing and CMA.@*CONCLUSION@#The IRF6: c.742G>T (p.G248C) heterozygous variant probably underlay the VWS in this pedigree. Above finding has also enabled reproductive guidance for the proband.

Chromosomal microarray analysis in prenatal diagnosis of women with isolated adverse pregnancy history / 中华围产医学杂志

Objective:To investigate the abnormal results of chromosomal microarray analysis (CMA) in the subsequent pregnancy of women with adverse pregnancy history, and explore the applicability of CMA in women with different genetic etiology.Methods:Out of 5 563 pregnant women who received CMA test in Nanjing Drum Tower Hospital during June 2014 and July 2020, 169 cases that underwent prenatal diagnosis due to isolated adverse pregnancy history were retrospectively collected in this study. All the participants were divided into three groups based on the etiology type of probands, genetic origin and expected CMA outcome: high-risk group ( n=19, including 11 cases with inherited pathogenic copy number variations and eight cases with inherited chromosomal abnormalities), low-risk group ( n=113, including six cases with negative whole exome sequencing and/or CMA findings, 31 cases with confirmed monogenic disease, 47 cases with de novo pathogenic copy number variations and 29 cases with de novo chromosomal abnormalities), and unknown risk group ( n=40, none of the cases underwent genetic testing). Descriptive statistical analysis was used to summarize the abnormal detection of each group. Results:There were 169 mothers with 172 fetuses finally enrolled, including two twins and one woman with two singleton pregnancies. A total of nine cases of abnormal fetuses were detected by CMA, accounting for 5.2% (9/172). Among them, eight were in the high-risk group, which were all caused by parental abnormalities, and one case in the low-risk group was detected with a de novo 22q11.22q11.23 microduplication, which was arr[GRCh37]22q11.22q11.23(22,997,928-25,002,659)×3. No abnormality was detected in the 40 patients of unknown risk group. Conclusions:Clarifying the etiology of isolated adverse pregnancy history is crucial to the rational application of CMA. Monogenic disease, unknown cause or negative finding of CMA in probands may not be an indication for prenatal diagnosis of CMA.

Genetic analysis and prenatal diagnosis of a pedigree with oculo-facio-cardio-dental syndrome: a case report and literature review / 中华围产医学杂志

Objective:To analyze the clinical phenotypes and prenatal diagnosis of a pedigree with oculo-facio-cardio-dental (OFCD) syndrome.Methods:A pregnant woman at 17 gestational weeks was admitted to the Nanjing Drum Tower Hospital, the Affiliated Hospital of Nanjing University Medical School in 2017 for genetic counseling. Genetic tests as performed for the proband (the pregnant woman), her husband, and the induced fetus of last pregnancy genetic test and the detected variants were analyzed and verified by chromosomal microarray analysis (CMA), multiplex ligation-dependent probe amplification (MLPA) and quantitative real time-polymerase chain reaction (Q-PCR). The detection platform established by MLPA and Q-PCR technology was used to perform prenatal diagnosis of the present pregnancy. Other family members were screened for BCOR gene mutation. Related mutation types were retrieved from ClinVar database with term of " BCOR", and related literature from CNKI and PubMed with terms of "OFCD syndrome", " BCOR gene", and "oculo facio cardiac dental syndrome" to summarize the clinical manifestations, mutation type and pathogenesis of this disease. Results:The proband has congenital cataracts, long face, congenital atrial septal defect, and severe dental malformations, which were consistent with the clinical features of OFCD syndrome. WES suggested that the proband and her induced fetus were suspected of having a large submicroscopic deletion of the exons of BCOR gene, which was confirmed by CMA, MLPA and Q-PCR, with a 105 kb deletion containing BCOR exons 1-15. The amniotic fluid genetic analysis of the present pregnancy showed that the fetus has a normal female karyotype, and did not carry the same BCOR gene copy number abnormality as the proband. The child grew and normally developed without any characteristic manifestations of OFCD syndrome during follow-up. Other families of the proband did not show clinical features of OFCD syndrone, and no BCOR gene copy number abnormality was detected. A total of 35 cases of BCOR gene mutation types related to OFCD syndrome were retrieved from ClinVar database. The data analysis revealed that the differences in clinical manifestations between Lenz microphthalmos syndrome and OFCD syndrome were mainly caused by different mutation types of BCOR gene. Among the 90 retrieved cases of OFCD syndrome obtained through literature, only one case was reported in China. Analysis of these 90 cases showed that the characteristic manifestations of OFCD syndrome, involving the eye, face, heart, teeth, and skeletal system. OFCD syndrome were confirmed in the proband and her induced fetus according to the clinical manifestation and the mutation type of BCOR gene. Conclusions:The clinical manifestations of OFCD syndrome are complicated, caused by various mutation types of BCOR. Systematic molecular genetic technology can be effectively applied for gene and prenatal diagnosis of OFCD syndrome.

Genetic study of a pedigree affected with oculodentodigital dysplasia / 中华医学遗传学杂志

Objective@#To explore the genetic basis of a pedigree affected with oculodentodigital dysplasia.@*Methods@#Genomic DNA was extracted from peripheral blood or amniotic fluid samples derived from the pedigree. Exon 2 of the GJA1 gene was amplified for sequencing.@*Results@#Two pedigree members were found to carry heterozygous missense variation of the GJA1 gene, c. 221A>C (p.H74P).@*Conclusion@#The missense c. 221A>C variation of the GJA1 gene probably underlies the oculodentodigital dysplasia in this pedigree.

Application of chromosomal microarray analysis in prenatal diagnosis of fetuses with increased nuchal translucency / 中华医学遗传学杂志

Objective@#To determine the frequency of chromosomal abnormalities and outcome of pregnancy for fetuses with increased nuchal translucency (NT).@*Methods@#Between July 2014 and February 2018, 247 fetuses with increased NT (>95th centile)were analyzed by chromosome microarray analysis (CMA). The fetuses were divided into ones with isolated increased NT(168 cases), increased NT with cystic hygroma(20 cases), increased NT with edema(12 cases) or increased NT with other abnormalities(47 cases). All couples were followed up by telephone calls.@*Results@#The rate of chromosomal abnormalities was 31.6% (78/247), which included 66 cases with chromosomal aneuploidies and 12 with copy number variants(CNVs). CNVs accounted for 31.4% (11/35) of total abnormalities among fetuses with isolated increased NT, whilst only 2.3% (1/43)of the total abnormalities among fetuses with non-isolated increased NT. Three fetuses with a normal CMA result had mental and physical retardation. Two of them were diagnosed with single gene disorders by whole exome sequencing.@*Conclusion@#CMA can detect more chromosomal microdeletion/microduplications among fetuses with isolated increased NT. Furthermore, fetuses with increased NT and anegative CMA result during pregnancy cannot exclude all adverse outcomes.

Genetic study of a pedigree affected with oculodentodigital dysplasia / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis of a pedigree affected with oculodentodigital dysplasia.@*METHODS@#Genomic DNA was extracted from peripheral blood or amniotic fluid samples derived from the pedigree. Exon 2 of the GJA1 gene was amplified for sequencing.@*RESULTS@#Two pedigree members were found to carry heterozygous missense variation of the GJA1 gene, c.221A>C (p.H74P).@*CONCLUSION@#The missense c.221A>C variation of the GJA1 gene probably underlies the oculodentodigital dysplasia in this pedigree.

Application of chromosomal microarray analysis in prenatal diagnosis of fetuses with increased nuchal translucency / 中华医学遗传学杂志

OBJECTIVE@#To determine the frequency of chromosomal abnormalities and outcome of pregnancy for fetuses with increased nuchal translucency (NT).@*METHODS@#Between July 2014 and February 2018, 247 fetuses with increased NT (>95th centile)were analyzed by chromosome microarray analysis (CMA). The fetuses were divided into ones with isolated increased NT (168 cases), increased NT with cystic hygroma (20 cases), increased NT with edema (12 cases) or increased NT with other abnormalities (47 cases). All couples were followed up by telephone calls.@*RESULTS@#The rate of chromosomal abnormalities was 31.6% (78/247), which included 66 cases with chromosomal aneuploidies and 12 with copy number variants (CNVs). CNVs accounted for 31.4% (11/35) of total abnormalities among fetuses with isolated increased NT, whilst only 2.3% (1/43) of the total abnormalities among fetuses with non-isolated increased NT. Three fetuses with a normal CMA result had mental and physical retardation. Two of them were diagnosed with single gene disorders by whole exome sequencing.@*CONCLUSION@#CMA can detect more chromosomal microdeletion/microduplications among fetuses with isolated increased NT. Furthermore, fetuses with increased NT and anegative CMA result during pregnancy cannot exclude all adverse outcomes.

Mutation analysis for a methylmalonic acidemia pedigree without proband by high-throughput sequencing / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To apply high-throughput sequencing for the detection of potential mutation in a methylmalonic academia pedigree for which no proband was available.</p><p><b>METHODS</b>For a couple who had previously given birth to an affected child, 14 genes were re-sequenced by high-throughput sequencing. Suspected mutations were validated by Sanger sequencing. Specific mutations were tested for amniotic fluid sample from the fetus.</p><p><b>RESULTS</b>High-throughput sequencing suggested that the husband has carried a heterozygous mutation of the MUT gene (Exon 3: c.729_730insTT; p.Asp244Leufs*39), while the wife also carried a heterozygous mutation of the MUT gene (Exon 5: c.914T>C; p.Leu305Ser). Both mutations were confirmed by Sanger sequencing. Testing of amniotic sample suggested that the fetus has carried neither mutation. Follow-up has found no sign of methylmalonic academia in the neonate.</p><p><b>CONCLUSION</b>High-throughput sequencing is a sensitive method to screen a bunch of genes in a single test. For autosomal recessive diseases, when no proband is available, carrier testing for both parents with high-throughput sequencing can provide an alternative approach, though great caution should be taken in the setting of prenatal diagnosis.</p>

Chromosomal microarray analysis of 85 fetus with growth restriction / 中华围产医学杂志

Objective To investigate the clinical value of chromosomal microarray analysis (CMA) in identifying the genetic etiology of fetal growth restriction (FGR).Methods Eighty-five FGR cases were recruited from Nanjing Drum Tower Hospital,the Affiliated Hospital of Nanjing University Medical School from January 2014 to October 2016.Samples ofamniotic fluid (n=74),skin tissues from aborted fetuses (n=9),umbilical cord blood (n=1) and peripheral blood from a premature infant (n=1) were collected.Affymetrix CytoScan 750K Array was used to detect copy number variation (CNV) in fetal samples.Microarray analysis.or fluorescence quantitative polymerase chain reaction was further recommended for the parents if fetal CMA result was variants of unknown significance (VOUS).Karyotype analysis of umbilical cord blood was further recommended if fetal CMA result was chromosome mosaicism.Chromosome analysis of peripheral blood was further recommended for the parents if fetal CMA result of a fetus was submicroscopic CNVs.Adjusted Chi-square test was used as the statistical method.Results CMA was successful in all samples in identifying chromosomal abnormalities.Among the 36 isolated FGR cases (42.4％,36/85),CMA identified in four cases of chromosome imbalance recombination and four cases of VOUS,and the rest 28 cases were normal.Besides,no CNV was detected.Among the other 49 FGR cases (57.6％,49/85) with ultrasound abnormalities,there were five cases of VOUS,and five cases of chromosome imbalance recombination and nine cases of CNVs.No significance difference in the detection rate of chromosome imbalance recombination was observed between the isolated and non-isolated FGR groups [11％ (4/36) vs 10％ (5/49),adjusted x2=0.000,P＞0.999].Parents of six cases of VOUS were further examined and the same variants was found in either one.One case of sex chromosome mosaicism was validated by cord blood karyotyping.One case of chromosome imbalance recombination was due to the paternal balanced translocation.Conclusions CMA is helpful in detecting the chromosome imbalance recombination in FGR cases.

Effects of galectin-3 on growth, migration and inflammation of endothe-lial cells / 中国病理生理杂志

AIM:To explore the effects of galectin-3 (GAL-3) on the viability, migration and inflammation of human umbilical vein endothelial cells (HUVECs) and the mechanisms.METHODS:The HUVECs were cultured in vitro and treated with GAL-3 recombinant protein at 2 mg/L or GAL-3 short hairpin RNA (shRNA).The HUVECs were divided into normal group, recombinant GAL-3 group, shControl group and GAL-3-shRNA group.The mRNA expression of GAL-3, monocyte chemotactic protein (MCP)-1, IL-6, matrix metalloproteinase (MMP)-9 and cyclin D1 was detected by real-time quantitative PCR,and the protein expression of GAL-3, IL-6 and MCP-1 was detected by Western blot.The secretion levels of MCP-1 and IL-6 in the culture medium were measured by ELISA.The viability and the ability of migration of the HUVECs were examined by CCK-8 assay and wound healing assay.The protein levels of heat shock protein 90 (HSP90), ERK1/2, p-ERK1/2, JNK and p-JNK were determined by Western blot.RESULTS:The expression of GAL-3, MCP-1 and IL-6 at mRNA and protein levels, the mRNA expression of MMP-9 and cyclin D1, and the secretion levels of MCP-1 and IL-6 in the culture medium were significantly higher than those in normal group (P<0.05) after the HUVECs were treated with GAL-3 recombinant protein.However, these molecules mentioned above in GAL-3-shRNA group were significantly lower than those in normal group and negative control group (P<0.05).Compared with normal group, the viability and migration ability of the HUVECs in recombinant GAL-3 group were significantly increased, but the viability and migration ability of the HUVECs in GAL-3-shRNA group were lower than those in normal group and shControl group (P<0.05).In addition, the protein levels of p-ERK1/2 and HSP90 in recombinant GAL-3 group were higher than those in normal group (P<0.05), but those in GAL-3-shRNA group were lower than those in normal group and shControl group (P<0.05).The protein level of p-JNK was not oviously changed among the 4 groups.CONCLUSION:GAL-3 is involved in regulating the cell growth, migration and the release of inflammatory cytokines in vascular endothelial cells, which may be mediated by HSP90-ERK1/2 signaling pathway.

Malaria endemic situation and comprehensive prevention and control mea-sures in Caoxian County / 中国血吸虫病防治杂志

Objective To understand the malaria endemic characteristics and control measures in Caoxian County,Shan?dong Province,so as to summarize the experiences of malaria elimination. Methods The data of malaria endemic situation and control measures in Caoxian County from 1953 to 2014 were collected and descriptively analyzed,and the control effectiveness was evaluated. Results The incidence of malaria reduced from 13.25%in 1970 to 0.33%in 1983,and no malaria case was found in 1986. The goal of basic malaria elimination was achieved. The sporadic malaria infections were found from 2006 to 2010,and three imported malaria cases were found in Caoxian County from 2011 to 2014. Conclusion The effect of compre?hensive prevention and control measures taken in Caoxian County is significant,and the goal of malaria elimination has been reached. The imported malaria and secondary cases are future focuses of malaria control work.

Study on osteogenic/chondrogenic differentiation potential of cartilage endplate derived stem cells in vivo / 局解手术学杂志

Objective To observe the osteogenic and/or chondrogenic differentiation potential of cartilage endplate derived stem cells ( CESCs) .Methods The cartilage endplate ( CEP) was obtained from 5 patients who underwent posterior discectomy procedure for the lumber degenerative disease.The agarose culture was used to select CESCs.The expanded CESCs were injected into the hydroxyapatite ceram-ic(HA) and incubated for 7 days.After that,the HA contained CESCs were implanted into the subcutaneous of nude mouse for 7 weeks.Then the HA was obtained and sliced for the specific stain and immunohistochemisty.The expanded CESCs were injected into the degenerated in-tervertebral disk of the rabbit for 16 weeks.Then MRI was used to detect the repair of the degenerated disc.Results The metachromasia of toluidin blue indicated that cartilage matrix was formed in the vacant space of HA.Mallory stain indicated that there was bone matrix was formed.The results of immunohistochemistry indicated that there was more collageⅠand collageⅩ,and a small quantity of collage Ⅱ.MRI results indicated that the CESCs could be induced into the chondrocyte-like cells in the rabbit intervertebral disk.Conclusion The CESCs could be induced into the bone/cartilage in vivo.

Effect of different routes of keyhole limpet hemocyanin immunization on the T cell dependent antibody response in mice / 中国实验动物学报

Objective To study the influence of different routes of keyhole limpet hemocyanin ( KLH) immuniza-tion on the T-cell-dependent antibody response in mice.Methods SPF Kunming mice were divided into four groups: the intravenous injection group, subcutaneous injection group, intraperitoneal injection group and control group.Each mouse was injected 200 μg KLH intravenously, subcutaneously or intraperitoneally daily for consecutive 10 days, respectively. Mice in the control group were given solvent injection only.Serum concentration of IgG stimulated by KLH antigen was measured 7 days after the last dosing.Spleen was isolated to calculate the organ coefficient and examined by pathology u-sing hematoxylin and eosin staining.Results Intravenously, subcutaneously and intraperitoneally administered KLH stimu-lated the generation of secondary lymphoid follicles and germinal center to varying degrees, B cell apoptosis, increased a-mount of cells in the marginal zone and other pathological changes were observed in the spleen.Intravenous and intraperito-neal administration of KLH led to more pronounced pathological changes compared with that in the subcutaneous injection group.All of the three administration routes of KLH induced generation of IgG antibody, significantly higher than that in the control group (P<0.05).Intravenous injection of KLH generated the highest concentration of IgG and organ coefficient among the three administration routes ( P<0.05) .Conclusions Different immunization routes do affect the production of IgG antibody, organ coefficient and pathological changes in the spleen, and these differences should be taken into consider-ation when analyzing the T cell dependent antibody response in mice.

Comparison of coagulation data measured with domestic produced and imported coagulation testing solutions on SD rat / 中国比较医学杂志

Objective To compare coagulation data measured with domestic produced and imported coagulation testing solutions on SD rat and human by testing prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time(TT), fibrinogen(FIB).Methods Blood samples were obtained from SPF SD rat and human .Domestic produced and imported coagulation testing solutions were applied to test PT , APTT, TT, and FIB.Results Compared to rat data measured with imported coagulation testing solution , data measured with domestic produced coagulation testing solution of PT, APTT, FIB was significantly higher (P<0.05), while, data of TT was statistically lower(P<0.05), and there was no obvious difference in human blood coagulation .Conclusion The data measured with different coagulation testing solution varies on SD rat , so the laboratories are required to establish reference data according to different products .

A retrospective study of 282 cases of pemphigus and cases of pemphigoid / 中华皮肤科杂志

Objective To analyze the diagnosis and therapy of pemphigus and pemphigoid.Methods Clinical data were retrospectively analysed for 648 patients with pemphigus or pemphigoid admitted in Huashan Hospital,Fudan University from 2003 to 2007.Results The mean age of onset of pemphigus was younger than that of pemphigoid(47.0±16.9 years vs 65.1±13.9 years.P＜0.001).Of 175 patients with pemphigus confirmed by direct immunoflorescence(DIF),100%were positive for IgG,and 92.0%for C3,while out of 223 patients with pemphigoid,51.12%were positive for IgG,99.1%for C3.For pemphigus,the consistency reached 68.8%among clinical diagnosis.pathological diagnosis and immunological(DIF)diagnosis,80.7%between pathological and immunological diagnosis.In the case of pemphigoid,the consistency was 62.8%among clinical,pathological and immunological diagnosis,and 78.1%between pathological and immunological diagnosis.Corticosteroids were primary treatment strategy for both pemphigus and pemphigoid.and prednisone of 0.5-1.5 mg per kilogram bodyweight per day could control the condition of most patients.Conclusions DIF could be used as an important diagnostic means for patients with pemphigus or pemphigoid which can not be confirmed by clinical manifestation or pathology.In primary hospitals,IgG and C3 are recommended for DIF testing when pemphigus is suspected,and C3 is recommended for suspected pemphigoid.